Methods for the administration of iloperidone

ABSTRACT

The present invention relates to methods for the identification of genetic polymorphisms that may be associated with a risk for QT prolongation after treatment with iloperidone and related methods of administering iloperidone to patients with such polymorphisms.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuation of co-pending U.S. patent applicationSer. No. 14/847,784, filed Sep. 8, 2015, which is a continuation of U.S.patent application Ser. No. 14/044,183, filed Oct. 2, 2013 (nowabandoned), which is a continuation of U.S. patent application Ser. No.12/208,027, filed Sep. 10, 2008 (now abandoned), which is acontinuation-in-part of U.S. patent application Ser. No. 11/576,178,filed Mar. 28, 2007 (now U.S. Pat. No. 8,586,610, issued Nov. 19, 2013),which is a 35 U.S.C. § 371 national stage entry of International PatentApplication No. PCT/US2005/035526, filed Sep. 30, 2005, which claims thebenefit of U.S. Provisional Patent Application No. 60/614,798, filedSep. 30, 2004. Each of the foregoing patent applications is incorporatedherein as though fully set forth.

SEQUENCE LISTING

The sequence listing contained in the electronic file entitled“VAND-0002-US-CIP-CON3_SequenceListing.txt,” created May 7, 2019 andcomprising 4 KB, is hereby incorporated herein by reference.

BACKGROUND OF THE INVENTION

Several genes associated with drug metabolism have been found to bepolymorphic. As a result, the abilities of individual patients tometabolize a particular drug may vary greatly. This can proveproblematic or dangerous where an increased concentration of anon-metabolized drug or its metabolites is capable of producing unwantedphysiological effects.

The cytochrome P450 2D6 gene (CYP2D6), located on chromosome 22, encodesthe Phase I drug metabolizing enzyme debrisoquine hydroxylase. A largenumber of drugs are known to be metabolized by debrisoquine hydroxylase,including many common central nervous system and cardiovascular drugs.One such drug is iloperidone(1-[4-[3-[4-(6-fluoro-1,2-benzisoxazol-3-yl)-1-piperidinyl]propoxy]-3-methoxyphenyl]ethanone).Iloperidone and methods for its production and use as an antipsychoticand analgesic are described in U.S. Pat. No. 5,364,866 to Strupczewskiet al. The diseases and disorders that can be treated by administrationof iloperidone include all forms of schizophrenia (i.e., paranoid,catatonic, disorganized, undifferentiated, and residual),schizoaffective disorders, bipolar mania/depression, cardiacarrhythmias, Tourette's Syndrome, brief psychotic disorder, delusionaldisorder, psychotic disorder NOS (not otherwise specified), psychoticdisorder due to a general medical condition, schizophreniform disorder,and substance-induced psychotic disorder. P88 is an active metabolite ofiloperidone. See, e.g., PCT WO2003020707, which is incorporated hereinby reference.

Among the unwanted physiological effects associated with an increasedconcentration of iloperidone or its metabolites is prolongation of theelectrocardiographic QT interval. Mutations in the CYP2D6 gene have beenassociated with a number of drug metabolism-related phenotypes. Theseinclude the ultra rapid metabolizer (UM), extensive metabolizer (EM),intermediate metabolizer (IM), and poor metabolizer (PM) phenotypes.Where a particular drug is capable of producing unwanted physiologicaleffects in its metabolized or non-metabolized forms, it is desirable todetermine whether a patient is a poor metabolizer of the drug prior toits administration.

A number of references are directed toward the identification of CYP2D6mutations and their corresponding phenotypes. For example, United StatesPatent Application Publication No. 2003/0083485 to Milos et al.describes a novel CYP2D6 variant associated with the PM phenotype andmethods for assessing whether an individual possesses the variant priorto the administration of a drug. United States Patent ApplicationPublication No. 2004/0072235 to Dawson describes a primer set useful inidentifying variants of the CYP2D6 gene. Similarly, United States PatentApplication Publication No. 2004/0091909 to Huang describes methods forscreening an individual for variants in the CYP2D6 gene and othercytochrome P450 genes and tailoring the individual's drug therapyaccording to his or her phenotypic profile. Finally, United StatesPatent Application Publication No. 2004/0096874 to Neville et al.describes methods for identifying cytochrome P450 variants.

SUMMARY OF THE INVENTION

The present invention comprises the discovery that treatment of apatient, who has lower CYP2D6 activity than a normal person, with a drugthat is pre-disposed to cause QT prolongation and is metabolized by theCYP2D6 enzyme, can be accomplishing more safely by administering a lowerdose of the drug than would be administered to a person who has normalCYP2D6 enzyme activity. Such drugs include, for example, dolasetron,paroxetine, venlafaxin, and iloperidone. Patients who have lower thannormal CYP2D6 activity are herein referred to as CYP2D6 PoorMetabolizers.

This invention also relates to methods for the identification of geneticpolymorphisms that may be associated with a risk for QT prolongationafter treatment with compounds metabolized by the CYP2D6 enzyme,particularly iloperidone or an active metabolite thereof or apharmaceutically acceptable salt of either (including, e.g., solvates,polymorphs, hydrates, and stereoisomers thereof), and related methods ofadministering these compounds to individuals with such polymorphisms.

The present invention describes an association between geneticpolymorphisms in the CYP2D6 locus, corresponding increases in theconcentrations of iloperidone or its metabolites, and the effect of suchincreases in concentrations on corrected QT (QTc) duration relative tobaseline. Any number of formulas may be employed to calculate the QTc,including, for example, the Fridericia formula (QTcF) and the Bazettformula (QTcB), among others. The present invention includes any suchformula or method for calculating a QTc.

A first aspect of the invention provides a method for treating a patientwith iloperidone or an active metabolite thereof or a pharmaceuticallyacceptable salt of either, comprising the steps of determining thepatient's CYP2D6 genotype and administering to the patient an effectiveamount of iloperidone or an active metabolite thereof or apharmaceutically acceptable salt of either based on the patient's CYP2D6genotype, such that patients who are CYP2D6 poor metabolizers receive alower dose than patients who are CYP2D6 normal metabolizers.

Another aspect of the invention provides a method for treating a patientwho is a CYP2D6 poor metabolizer with iloperidone or an activemetabolite thereof or a pharmaceutically acceptable salt of either,wherein the patient is administered a lower dosage than would be givento an individual who is not a CYP2D6 poor metabolizer.

Another aspect of the invention provides a method of treating a patientwith iloperidone or an active metabolite thereof or a pharmaceuticallyacceptable salt of either comprising the steps of determining whetherthe patient is being administered a CYP2D6 inhibitor and reducing thedosage of drug if the patient is being administered a CYP2D6 inhibitor.

Another aspect of the invention provides a method for determining apatient's CYP2D6 phenotype comprising the steps of administering to thepatient a quantity of iloperidone or an active metabolite thereof or apharmaceutically acceptable salt of either, determining a firstconcentration of at least one of iloperidone and an iloperidonemetabolite in the patient's blood, administering to the patient at leastone CYP2D6 inhibitor, determining a second concentration of at least oneof iloperidone and an iloperidone metabolite in the patient's blood, andcomparing the first and second concentrations.

Another aspect of the invention provides a method for determiningwhether a patient is at risk for prolongation of his or her QTc intervaldue to iloperidone administration comprising the step of: determining apatient's CYP2D6 metabolizer status by either determining the patient'sCYP2D6 genotype or CYP2D6 phenotype. In the case that a patient isdetermined to be at risk for prolongation of his or her QTc interval,the dose of iloperidone administered to the patient may be reduced.

Another aspect of the invention provides a method of administeringiloperidone or an active metabolite thereof, or a pharmaceuticallyacceptable salt of either, for the treatment of a disease or disorder ina human patient comprising the steps of determining the activity of thepatient's CYP2D6 enzyme on at least one of iloperidone and itsmetabolites relative to the activity of a wild type CYP2D6 enzyme andreducing the dose of at least one of iloperidone and itspharmaceutically acceptable salts if the patient's CYP2D6 enzymeactivity is less than that of the wild type CYP2D6.

Another aspect of the invention relates to modifying the dose and/orfrequency of dosing with iloperidone or a pharmaceutically acceptablesalt thereof based on the P88:P95 ratio and/or the (P88+iloperidone):P95ratio in a blood sample of a patient being treated with iloperidone orP88, especially patients susceptible to QT prolongation or to harmfuleffects associated with QT prolongation.

Another aspect of the invention provides a kit for use in determining aCYP2D6 genotype of an individual, comprising a detection device, asampling device, and instructions for use of the kit.

Another aspect of the invention provides a kit for use in determining aCYP2D6 phenotype of an individual, comprising a detection device, acollection device, and instructions for use of the kit.

Another aspect of the invention provides a kit for use in determining atleast one of a P88 to P95 ratio and a P88 and iloperidone to P95 ratioin an individual, comprising a detection device, a collection device,and instructions for use of the kit.

Yet another aspect of the invention provides a method forcommercializing a pharmaceutical composition comprising at least one ofiloperidone, a pharmaceutically acceptable salt of iloperidone, anactive metabolite of iloperidone, and a pharmaceutically acceptable saltof an active metabolite of iloperidone, said method comprising:obtaining regulatory approval of the composition by providing data to aregulatory agency demonstrating that the composition is effective intreating humans when administered in accordance with instructions todetermine whether or not a patient is a CYP2D6 poor metabolizer prior todetermining what dose to administer to the patient; and disseminatinginformation concerning the use of such composition in such manner toprescribers or patients or both.

The foregoing and other features of the invention will be apparent fromthe following more particular description of embodiments of theinvention.

DETAILED DESCRIPTION OF THE INVENTION

Iloperidone is a benzisoxazole-piperidinyl derivative, currently indevelopment for the treatment of CNS disorders. Data fromplacebo-controlled Phase m studies of iloperidone showed a Fridericiacorrection of QT duration (QTcF) increase of 0.1 to 8.5 msec at doses of4-24 mg, when comparing a single ECG at baseline to a single ECG atendpoint. At lower doses of iloperidone (4 mg-16 mg) QTcF prolongationwas minimal (0.1-5 msec). In the most recent study, a greaterprolongation was observed when higher doses of iloperidone (20-24mg/day) were studied. The mean change in the QTcF at doses 20-24 mg/daywas 8.5 msec, and 4.6 msec in the 12-16 mg/day dose range in this study.These data suggest that treatment with iloperidone can be associatedwith prolongation of the QT interval similar to other drugs in thisclass, and that the effect may be dose sensitive in the clinical doserange.

The research leading to the present invention was designed to examinethe effect of different doses of iloperidone relative to the effect ofziprasidone and quetiapine on QTc duration under carefully controlledconditions. To further evaluate the possible relationship betweenexposure to iloperidone and the comparators to QTc duration,reassessment after pharmacological inhibition of the principle metabolicpathways for each drug, under steady-state conditions, was also planned.

Example 1

Blood samples for pharmacogenetic analysis were collected at screening.Two polymorphisms previously associated with poor metabolizing statuswere genotyped in the CYP2D6 locus, and 251 genotypes were collected.The individual genotypes were studied for detection of associationbetween genotype class and concentrations of iloperidone and itsmetabolites P88 and P95. The functional effect of the polymorphisms wasalso evaluated by analyzing the effect of the addition of the CYP2D6inhibitor paroxetine on the concentrations of the parent drug and itsmetabolites.

The research leading to the present invention identified a significantassociation between CYP2D6 genotype and concentrations of P88 before theaddition of inhibitors as well as the effect of this association on QTcprolongation.

Iloperidone is a substrate for two P450 enzymes; CYP2D6 and CYP3A4. Mostmetabolic clearance of iloperidone depends on these two enzymes. CYP2D6catalyzes hydroxylation of the pendant acetyl group to form metaboliteP94, which is converted to P95 after some additional reactions. Additionof the CYP2D6 inhibitor fluoxetine, along with iloperidone resulted inincreases of the area under the curve (AUC) for iloperidone and P88 of131% and 119% respectively. Addition of the CYP3A4 inhibitorketoconazole in interaction studies resulted in a 38-58% increase in theconcentrations of iloperidone and its main metabolites P88 and P95. P88has a pharmacological profile including affinity for the HERG channelsimilar to that of iloperidone. P95 is less lipophilic and is dissimilarin its binding profile compared to iloperidone, including having verylow affinity for the HERG channel. For these reasons P95 is regarded asbeing pharmacologically inactive.

The addition of metabolic inhibitors in this study therefore allowed foran evaluation of the effect of increasing blood-concentration ofiloperidone and/or its metabolites on QT duration. More specifically,this study allowed for an evaluation of the effect of iloperidone on QTcbefore and after the addition of the CYP2D6 inhibitor, paroxetine, aswell as before and after the addition of the CYP3A4 inhibitor,ketoconazole.

The CYP2D6 gene is highly polymorphic, with more than 70 allelicvariants described so far. See, e.g.,www.imm.ki.se/CYPalleles/CYP2D6.htm. Most embodiments of the presentinvention concern the two most common polymorphisms within the CYP2D6gene in Caucasian populations, CYP2D6G)846A and CYP2D6P34S (alsoreferred to as CYP2D6C100T). These polymorphisms correspond tonucleotides 3465 and 1719, respectively, in GenBank sequence M33388.1(GI:181303). The CYP2D6P34S/CYP2D6C100T polymorphism also corresponds tonucleotide 100 in GenBank mRNA sequence M20403.1 (GI:181349).

The CYP2D6G)846A polymorphism (known as the CYP2D6*4 alleles,encompassing *4A, *4B, *4C, *4D, *4E, *4F, *4G, *4H, *4J, *4K, and *4L)represents a G to A transition at the junction between intron 3 and exon4, shifting the splice junction by one base pair, resulting inframeshift and premature termination of the protein (Kagimoto 1990,Gough 1990, Hanioka 1990). The CYP2D6P34S/CYP2D6C100T polymorphism(known as the CYP2D6*10 and CYP2D6*14 alleles) represents a C to Tchange that results in the substitution of a Proline at position 34 bySerine (Yokota 1993, Johansson 1994). Both of these polymorphisms havebeen associated with reduced enzymatic activity for different substrates(Johansson 1994, Dahl 1995, Jaanson 2002, see also review by Bertilsson2002)

Methods

A. Samples

128 individuals consented to the pharmacogenetic study. Blood sampleswere collected according to the pharmacogenetics protocol and after theconsent of patients. The DNA was extracted from whole blood by Covanceusing the PUREGENE DNA isolation kit (D-50K).

The 128 individuals that participated were a good representation of thetotal sample of 165 individuals that participated in the trial. 22 of 29total were from the iloperidone 8 mg bid group, 30 of 34 were from theiloperidone 12 mg bid group, 22 of 31 from the 24 mg qd group, 3 of 5 ofthe risperidone group, 28 of 33 of the ziprazidone group, and 23 of 33of the quetiapine group.

B. Genotyping

Genotypes for the CYP2D6G1846A polymorphism were ascertained for 123 ofthe 128 consenting individuals, while genotypes for the CYP2D6C100Tpolymorphism were identified for all 128 participants. Genotyping wasperformed on amplified DNA fragments. The CYP2D6 genomic region wasamplified using a triplex PCR strategy (Neville 2002). In brief, primersused were:

Exons 1 & 2 SEQ. ID. 1, 2D6L1F1: CTGGGCTGGGAGCAGCCTCSEQ. ID. 2, 2D6L1R1: CACTCGCTGGCCTGTTTCATGTC Exons 3, 4, 5 & 6SEQ. ID. 3, 2D6L2F: CTGGAATCCGGTGTCGAAGTGG SEQ. ID. 4, 2D6L2R2:CTCGGCCCCTGCACTGTTTC Exons 7,8 & 9 SEQ. ID. 5, 2D6L3F:GAGGCAAGAAGGAGTGTCAGGG SEQ. ID. 6, 2D6L3R5B: AGTCCTGTGGTGAGGTGACGAGG

Amplification was performed on 40-100 ng of genomic DNA using a GC-richPCR kit (Roche Diagnostics, Mannheim, Germany) according to themanufacturer's recommendations. Thermocycling conditions were asfollows: initial denaturation (3 min 95° C.), 10 cycles of 30 s ofdenaturation (30 s at 95° C.), annealing (30 s at 66° C.), andextension, (60 s at 72° C.) followed by 22 cycles: 30 s at 95° C., 30 sat 66° C., 60 s+5 s/cycle at 72° C. A final extension followed (7 min at72° C.).

Third Wave Technologies, Inc (Madison, Wis.) developed the probe setsfor genotyping. Genotyping was performed on PCR products using theInvader® assay (Lyamichev 1999) (Third Wave Technologies, Inc) accordingto the manufacturer's recommendations.

The genotypes of individuals distributed among the three iloperidonegroups were not significantly different (Table 1A and 1B).

TABLE 1A Genotype frequencies by iloperidone dose class for CYP2D6C100TIloperidone Genotype dose group CC CT TT Total Ilo 8 mg bid  19^(a) 2 122 Ilo 12 mg bid 23 6 1 30 Ilo 24 mg qd 15 6 1 22 Total 57 14 3 74^(a)number of individuals

TABLE 1B Genotype frequencies by iloperidone dose class for CYP2D6G1846AIloperidone Genotype dose group AA AG GG Total Ilo 8 mg bid 0 3 17 20Ilo 12 mg bid 1 6 23 30 Ilo 24 mg qd 1 5 15 21 Total 2 14 55 71

C. Statistical Analysis

The genotype effect of the two CYP2D6 polymorphisms on period 1concentrations was evaluated using the following ANOVA model.Concentrations of iloperidone, P88, and P95 at Period 1, withoutinhibitor, at the time at which maximum blood concentration of theparent compound or metabolite was reached (Tmax) were used as thedependent variable, the genotypes of each polymorphism as classes andthe treatment as a covariate. In order to adjust for treatment effectsafter the single dose of iloperidone, the 8 mg bid was coded as 8, the12 mg bid as 12 and the 24 mg qd as 24.

The function of these polymorphisms on the degree of inhibition of theCYP2D6 enzyme was calculated from the ratio of concentrations of P88 andP95 in period 2, after the addition of the inhibitor of CYP2D6. Theconcentrations of iloperidone and/or its metabolites (e.g., P88 and P95)may be determined in period 1 and/or period 2 by any known orlater-developed method or device, including titration.

Results and Discussion

In order to understand the functional significance of the two CYP2D6polymorphisms on the activity of the enzyme, the association of thevarious genotypes with the relative concentrations of the metabolitesP88 and P95 were examined. It is known that P88 is degraded by CYP2D6and that CYP2D6 is involved in the synthesis of P95. The relativeamounts of P88 and P95 would therefore be controlled by the activity ofthe CYP2D6 enzyme. The ratio of P88/P95 was calculated before inhibitionin Period 1 and at the Tmax of the two metabolites, as well as the ratioof P88/P95 in Period 2 after the addition of the CYP2D6 inhibitorparoxetine. In individuals with the wild type enzyme the concentrationof P88 is expected to increase in Period 2, while in the same period theconcentration of P95 is expected to decline.

For Period 1 the mean P88/P95 ratio among the 91 iloperidone treatedpatients was equal to 1.0 with a range from 0.14 to 8.19. Among the sameindividuals for Period 2 the mean ratio was 2.4 with a range from 0.5 to8.49. The mean ratio of the ratios Period 1/Period 2 was equal to 0.37with a range from 0.11 to 2.75.

Among the genotyped individuals the values were similar with means of 1;2.45; and 0.37 for Period 1, Period 2, and Period 1/Period 2respectively, indicating no sample bias. For polymorphism CYP2D6G)846Athe means were significantly different between the three-genotypeclasses AA, AG and GG. For AA the respective values were 6.1, 3.41, and1.89; for AG they were 2.4, 4.2, and 0.52; and for GG, 0.57, 1.94, and0.28 (Table 2).

TABLE 2 Ratios of P88, P95 concentrations according to genotype Popula-P88/P95 tion P88/P95 Period1 P88/P95 Period 2 (Period1/Period2) All 1.0(0.14-8.19) 2.45 (0.50-8.49) 0.37 (0.11-2.75) CYP2D6G1846A AA 6.1(3.96-8.19) 3.41 (2.96-3.87) 1.89 (1.0-2.75) AG 2.4 (0.44-7.0) 4.20(2.2-7.57) 0.52 (0.14-1.28) GG 0.57 (0.14-2.2) 1.94 (0.52-4.71) 0.28(0.11-0.61)

The differences between genotype classes were significant at thep<0.0001 level in ANOVA test. These data suggest that the AA classrepresent a CYP2D6 poor metabolizer as indicated by the high ratio ofP88/P95 in period 1 and the relatively small effect of the addition ofthe inhibitor in Period 2. The AG class seems to exhibit an intermediatephenotype between the poor metabolizer and the wild type with anapproximately 2-fold reduction of the CYP2D6 activity after the additionof the inhibitor, as indicated by the ratio of the ratios (Table 2).This analysis provides a phenotypic characterization of the CYP2D6G)846Apolymorphism as it relates to the metabolism of iloperidone.

Having established a functional role of this polymorphism, theconcentrations of P88 at Period 1 at the Tmax of P88 were calculated foreach genotype class. P88 concentrations were significantly (p<0.005)higher for the AA and AG classes as compared to the GG class for each ofthe three iloperidone dose groups (Table 3).

TABLE 3 P88 concentrations in Period 1 according to CYP2D6 genotypeGenotype N obs LSMeans P value AA 2 62.70 <0.0001 AG 14 31.40 GG 5521.03 TRT dose 0.0015 CYP2D6G1846A *TRT dose 0.0058

Although the number of individuals carrying the A allele is limited, theresults obtained in the study consistently suggest that individuals ofthe AA and AG class are expected to experience higher concentrations ofP88 at Tmax as compared with GG individuals. Similar results wereobtained with polymorphism CYP2D6C100T (Table 4 and 5).

TABLE 4 Ratios of P88, P95 concentrations according to genotype Popula-P88/P95 tion P88/P95 Period1 P88/P95 Period 2 (Period1/Period2) All 1.0(0.14-8.19) 2.45 (0.50-8.49) 0.37 (0.11-2.75) CYP2D6C100T CC 0.6(0.14-2.28) 1.93 (0.52-4.71) 0.27 (0.11-0.61) CT 2.2 (0.44-7.0) 4.14(2.2-7.57) 0.49 (0.14-1.28) TT 5.24 (3.56-8.19) 4.19 (2.96-5.74) 1.46(0.62-2.75)

TABLE 5 P88 concentrations in Period 1 according to CYP2D6 genotypeGenotype N obs LSMeans P value CC 57 21.03 CT 14 33.16 <0.0001 TT 351.00 TRT dose <0.0001 CYP2D6C100T *TRT dose 0.0015

This result is expected given the fact that this polymorphism is inalmost complete linkage disequilibrium with the CYP2D6G)846Apolymorphism.

In order to understand whether the difference in concentration of P88 atPeriod 1 Tmax was relevant to the increases in QTc after the addition ofthe inhibitors, the observed mean of P88 for the CYP2D6G)846A AG groupwas used to divide all individuals into two classes. The first includesindividuals with P88 concentrations at Period 3, after the addition ofboth inhibitors, of equal to or less than 34 ng/mL and the second classincludes individuals with P88 concentration greater than 34 ng/mL. Thetwo classes were then compared in regards to the QTc change frombaseline at Period 3. Using an ANOVA statistic for the first classP88>34 (n=55) the QTc mean change from baseline in Period 3 was 22.7msec and that for P88≤34 (n=12) the mean QTc for the same period was 7.7msec. The QTc changes from baseline for Period 1 and Period 2 accordingto genotype and iloperidone dose are given in Table 6 and 7.

TABLE 6 QTc change at Period 1 according to CYP2D6 genotype andiloperidone dose Iloperidone Dose Genotype 8 mg bid 12 mg bid 24 mg qdCYP2D6G1846A AA 17.7 (1)^(a) 38.4 (1) AG −0.8 (3) 5.8 (6) 19.0 (5) GG7.8 (17) 11.8 (23) 14.0 (14) CYP2D6C100T TT −8.4 (1) 17.7 (1) 38.4 (1)CT 2.9 (2) 5.8 (6) 19.0 (5) CC 7.8 (17) 11.8 (23) 9.5 (14) ^(a)number ofindividuals

TABLE 7 QTc change at Period 2 according to CYP2D6 genotype andiloperidone dose Iloperidone Dose Genotype 8 mg bid 12 mg bid 24 mg qdCYP2D6G1846A AA 25.0 (1) 28.4 (1) AG 8.1 (3) 8.7 (6) 20.6 (5) GG 11.7(18) 14.5 (21) 16.4 (15) CYP2D6C100T TT −0.7 (1) 25.0 (1) 28.4 (1) CT12.5 (2) 8.7 (6) 20.6 (5) CC 11.7 (16) 14.5 (21) 16.4 (15)

These results should be viewed with caution, however, since the numberof observations is small. If one was to focus on the iloperidone 24 mgqd, there is a trend for higher QTc among AA, and AG individuals forCYP2D6G)846A as compared to GG. This difference disappears after theaddition of the CYP2D6 inhibitor in Period 2.

These observations suggest that the differences in P88 concentrationsduring Period 1 between the different classes of genotypes may berelevant to QTc changes from baseline. Given the small number ofobservations and the unbalanced in regards to genotype design of thestudy, a confirmatory prospectively designed study may be requiredbefore any further interpretation of this data is warranted.

Notwithstanding these caveats, the results discussed above show thatpatients can be more safely treated with iloperidone if the dose ofiloperidone is adjusted based on the CYP2D6 genotype of each patient.For example, if a patient has a genotype which results in decreasedactivity of the CYP2D6 protein relative to the wild type CYP2D6, thenthe dose of iloperidone administered to such patient would be reducedto, for example, 75% or less, 50% or less, or 25% or less of the dosetypically administered to a patient having a CYP2D6 genotype thatresults in a CYP2D6 protein that has the same or substantially the sameenzymatic activity on P88 as the wild type CYP2D6 genotype/protein. Forexample, where the normal dosage of iloperidone or otherCYP2D6-metabolized compound administered to an individual is 24 mg perday, an individual with a genotype associated with decreased CYP2D6activity may receive a reduced dosage of 18, 12, or 6 mg per day.

Decreased CYP2D6 activity may be the result of other mutations,including those described at www.imm.ki.se/CYPalleles/CYP2D6.htm, whichis incorporated herein by reference. In particular, it is noted that theCYP2D6*2A mutation includes a CYP2D7 gene conversion in intron 1. Insome cases, the lower CYP2D6 activity in a CYP2D6 poor metabolizer maybe due to factors other than genotype. For example, a patient may beundergoing treatment with an agent, e.g., a drug that reduces CYP2D6activity.

QTc prolongation is correlated to the ratios of P88/P95 and(iloperidone+P88)/P95. The mean ratios among CYP2D6 extensivemetabolizers were 0.57 and 1.00, respectively. As shown above in Tables3 and 5, CYP2D6 poor metabolizers have elevated P88 levels compared toCYP2D6 extensive metabolizers.

As CYP2D6 poor metabolizers comprise approximately 15% of thepopulation, it was found that approximately 15% of those studiedexhibited a P88/P95 ratio greater than 2.0 while the remaining 85%exhibited P88/P95 ratios less than 2.0. Table 8 below shows the leastsquares mean change in QTc for each dosage group. While the results forsome groups are not statistically significant, they do indicate a trendsupporting the hypothesis that QTc prolongation is correlated to P88/P95ratio. Similar results were obtained when cutoff ratios of 3.0 and 4.0were analyzed, providing further support to the hypothesis that theextent of QTc prolongation a patient may experience after treatment canbe predicted by measuring P88 and P95 blood levels.

TABLE 8 Mean QTc Prolongation According to P88/P95 Ratio LSMean LSMeanLSMean LSMean LSMean QTc change QTc change QTc change QTc change QTcchange from Baseline P88/P95 from Baseline from Baseline from Baselinefrom Baseline All Treatment Ratio 8 mg bid 12 mg bid 8 + 12 mg bid 24 qdGroups <2 7.2 (n = 23) 8.7 (n = 31) 8.3 (n = 54) 13.9 (n = 24) 10.244 (n= 78) >2 21.3 (n = 5) 17.4 (n = 3) 18.3 (n = 8) 29.4 (n = 5) 21.111 (n =13) P value 0.0725 0.392 0.0815 0.0329 0.0131

Similar results were observed when considering QTc correlation to the(iloperidone+P88)/P95 ratio. Again, as approximately 15% of thepopulation are CYP2D6 poor metabolizers, it was found that approximately15% of those studied exhibited (iloperidone+P88)/P95 ratios greater than3.0 while the remaining 85% exhibited ratios less than 3.0. Table 9below shows the least squares mean change in QTc for each dosage group.While the results for some groups are not statistically significant,they do indicate a trend supporting the hypothesis that QTc prolongationis correlated to (iloperidone+P88)/P95 ratio. Indeed, when cutoff ratiosof 3 and higher were analyzed, similar results were obtained providingfurther support to the hypothesis that the extent of QTc prolongation apatient may experience after treatment can be predicted by measuringiloperidone, P88 and P95 blood levels.

TABLE 9 Mean QTc Prolongation According to (iloperidone + P88)/P95 RatioLSMean LSMean LSMean LSMean LSMean QTc change QTc change QTc change QTcchange QTc change from Baseline (ILO + P88)/ from Baseline from Baselinefrom Baseline from Baseline All Treatment P95 Ratio 8 mg bid 12 mg bid8 + 12 mg bid 24 qd Groups <3 7.2 (n = 23) 8.7 (n = 31) 8.3 (n = 54)14.4 (n = 24) 10.424 (n = 78) >3 21.3 (n = 5) 15.2 (n = 3) 17.3 (n = 8)30.5 (n = 5) 20.031 (n = 13) P value 0.0725 0.4223 0.0857 0.0522 0.0278

While the CYP2D6G)846A (AA or AG) genotype and the CYP2D6C100T (CT orTT) genotype are illustrated in this Example 1, the method of theinvention can employ other genotypes that result in decreased activityof the CYP2D6 protein on iloperidone and P88. It is within the skill ofthe art, based on the disclosure herein, to identify additional CYP2D6genotypes that result in decreased enzymatic activity on iloperidone andP88.

Example 2

A second study extended the pharmacogenomic assessment of iloperidoneresponse by genotyping additional CYP2D6 variants which lead to theproduction of a non-functional protein or reduced enzymatic activity.

Six of the variants have been shown to result in the absence of afunctional enzyme, either because of a deletion of the gene (as in theCYP2D6 *5 polymorphism), a frameshift (*3 and *6), a splicing error(*4), or a truncated or abnormal protein (*7 and *8). Five otherpolymorphisms were genotyped that resulted in the production of afunctional protein that was shown to have a significantly decreasedenzymatic activity on various compounds such as debrisoquine orsparteine (*9, *10, *17, *41), or only a modest reduction in activity(*2). The actual impact of these individual polymorphisms on the enzymevary from compound to compound, and the presence of several of them inthe same protein can further reduce the CYP2D6 activity.

Methods

A. Samples

From the 300 iloperidone-treated patients initially genotyped for theCYP2D6*4 and *10 variants (VP-VYV-683-3101), 222 remaining DNA sampleswere used for this extended pharmacogenomic analysis. One patient wasexcluded from the analysis due to inconsistent results for the CYP2D6allele *4 generated by Quest Diagnostics Central Laboratory(Collegeville, Pa.) and Cogenics (Morrisville, N.C.). Pharmacokineticdata of the (iloperidone+P88)/P95 ratio was available for 168 of thesepatients. QT measurement at Day 14 and 28 was available for 169 and 146patients respectively.

B. Genotyping

Eleven specific CYP2D6 polymorphisms were evaluated (Table 10).

TABLE 10 CYP2D6 polymorphisms Enzyme Allele DNA variations Effectactivity *1 Wild type Normal *2 2850C > T; 4180G > C R296C; S486T Normal(dx, d, s) *3 2549del 259Frameshift None (d, s) *4 100C > T; 1661G > C;1846G > A P34S; splicing None (d, s) defect *5 CYP2D6 deletion CYP2D6deleted None (d, s) *6 1707delT 118Frameshift None (d, s, dx) *7 2935A >C H324P None (s) *8 1661G > C; 1758G > T; 2850C > T; 4180G > C G169XNone (d, s) *9 2615 _(—) 2617delAAG K281del Decreased (b, d, s) *10100C > T; 1661G > C; 4180G > C P34S; S486T Decreased (d, s) *17 1023C >T; 1661G > C; 2850C > T; 4180G > C T107I; R296C; Decreased (d) S486T *41−1584C; −1235A > G; −740C > T; −678G > A; R296C; splicing Decreased (s)CYP2D7 gene conversion in intron 1; 1661G > C; 2850C > T; defect; S486T2988G > A; 4180G > C The DNA variations and their effects at the RNA orprotein level listed here are based on the description by the HumanCytochrome P450 (CTP) Allele Nomenclature Committee available at:http://www.cypalleles.ki.se/CYP2D6.htm. The in-vivo changes in enzymeactivity have been reported for bufuralol (b), debrisoquine (d),dextromethorphan (dx), or sparteine (s). The specific polymorphismsgenotyped in the study reported here are shown in bold.

The genotypes of the CYP2D6* 10 allele were generated by QuestDiagnostics Central Laboratory (Collegeville, Pa.); the genotypes of theCYP2D6 *2, *3, *5, *6, *7, *8, *9, *17 and *41 alleles were generated byCogenics (Morrisville, N.C.); and the genotypes of the CYP2D6 *4 allelewere obtained from Quest and also from Cogenics for a subset ofpatients.

The CYP2D6*2 allele is characterized by a series of mutations. In thisassay, the cytosine to thymine transition at position 2850, whichresults in an arginine to cysteine substitution at amino acid 296, wastested (Johansson et al., 1993; Wang et al. 1995). The first roundproduct from CYP2D6 multiplex PCR was amplified and the resultingproduct was digested with HhaI. The HhaI digestion resulted in 476, 372,247, 178, and 84 basepair fragments for wt/wt genotype; 550, 476, 372,247, 178, and 84 basepair fragments for *2/wt genotype; and 550, 476,247, and 84 basepair fragments for *2/*2 genotype. The PCR products weregel electrophoresed and photographed under ultraviolet light.

The presence of the CYP2D6 alleles *3, *4, *6, *7, and *8 was assayedusing multiplex PCR (Stuven et al. 1996). The CYP2D6 *3 allele resultsfrom a single base (adenine) deletion at nucleotide 2549 in exon 5(Buchert et al., 1993). The defect in the CYP2D6 *4 allele is due to aguanine to adenine transition in the last nucleotide (position 1846) ofintron 3 resulting in an aberrant 3′ splice recognition site (Hanioka etal. 1990). The CYP2D6 *6 allele results from a thymine deletion atposition 1707 in exon 3 resulting in a premature stop codon (Saxena etal. 1994). The CYP2D6 *7 allele results from an adenine to cytosinemissense mutation at position 2935 which results in a histidine toproline substitution at amino acid 324 in exon 6 leading to a total lossof enzyme function (Evert et al. 1994). The defect in the CYP2D6 *8allele is due to a guanine to thymine transition at position 1758resulting in the insertion of a premature stop codon (Stuven et al.1996).

The first round amplification generated a 1578 basepair productcontaining all five alleles. The 1578 basepair product served as thetemplate for a multiplex allele-specific assay to simultaneouslyidentify the five alleles. First round PCR template was added to twoseparate master mixes containing primers that recognize wild type ormutant alleles. These primers produce PCR products of 1394, 1010, 304,219, and 167 basepairs for *7, *3, *4, *8, and *6 alleles, respectively.As an internal control, the primers for *8 were reversed; that is, theprimer that recognizes the wild-type allele for *8 was present in themutant master mix and the primer for the mutant allele for *8 waspresent in the wild-type master mix. For wild-type genotypes (except for*8), PCR products appeared in the wild-type lanes while no PCR productswere observed in the mutant lane. For heterozygous genotypes, PCRproducts of the same fragment size appeared in both the wild-type andmutant lanes. For mutant genotypes (except for *8), PCR product appearedonly in the mutant lane. The PCR products were gel electrophoresed andphotographed under ultraviolet light.

The CYP2D6 *5 allele results from a complete deletion of the CYP2D6 gene(Gaedigk et al. 1991; Steen et al. 1995). A long-range PCR method wasused to identify a deletion of the CYP2D6 locus. Presence or absence andintensity of PCR products identified the wild-type, heterozygous mutant,or mutant alleles. The PCR products were gel electrophoresed andphotographed under ultraviolet light.

The CYP2D6 *9 mutation is a 3 basepair deletion at positions 2613-2615(Tyndale et al. 1991). This results in a deletion of lysine at aminoacid 281. The CYP2D6 *41 mutation is due to a guanine to adeninetransition at position 2988 (Raimundo et al. 2004). The first round ofamplification generated a 1578 basepair product containing the twoalleles. The 1578 basepair product served as the template for amultiplex allele-specific assay to simultaneously identify the twoalleles. First round PCR template was added to two separate master mixescontaining primers that recognize wild-type or mutant alleles. Theseprimers produced PCR products of 409, 593, and 780 basepairs for the *9wild-type, internal control, and *41 wild-type, respectively. Forwild-type genotypes, PCR products appeared in the wild-type lanes whileno PCR products were observed in the mutant lane. For heterozygousgenotypes, PCR products appeared in both the wild-type and mutant lanes.For mutant genotypes, PCR product appeared only in the mutant lane. ThePCR products were gel electrophoresed and photographed under ultravioletlight.

The CYP2D6 * 17 allele results from a cytosine to thymine base change atposition 1023 which results in a threonine to isoleucine substitution atamino acid 107 in exon 2 (Masimirembwa et al. 1996). The first round ofamplification generated a 369 basepair product containing the CYP2D6 *17allele. The first round PCR template was added to two separate mastermixes containing primers that recognize wild-type or mutant alleles aswell as an internal control. These primers produced PCR products of 235and 181 basepairs for the * 17 allele and internal control,respectively. For a wild-type genotype, both PCR products appeared inthe wild-type lanes while only the internal control PCR product wasobserved in the mutant lane. For heterozygous genotypes, both PCRproducts appeared in both the wild-type and mutant lanes. For a mutantgenotype, both PCR products appeared in the mutant lanes while only theinternal control PCR product is observed in the wild-type lane. The PCRproducts were gel electrophoresed and photographed under ultravioletlight.

C. Statistical Analysis

Analyses were performed on observed case data using an ANCOVA model withthe baseline value as a covariate for the change from baseline in QTc(Fridericia formula) and using an ANOVA model for iloperidone bloodexposure at Day 14 and Day 28. Linkage disequilibrium analysis wasperformed using Haploview v4.0 (Barrett et al, 2005).

Statistically significant associations were observed between the CYP2D6*4, CYP2D6 *5, CYP2D6 *10, and CYP2D6 *41 alleles and the iloperidoneblood exposure levels. The ratio of drug concentration[(iloperidone+P88)/P95] was increased with the presence ofnon-functional CYP2D6 alleles and of variants possibly associated withdecreased enzymatic activity. Furthermore, patients who carried at leastone non-functional CYP2D6 allele had a higher QTc prolongation after 14days of iloperidone treatment than those with two functional copies. ByDay 28, the QTcF prolongation was reduced but was still statisticallydifferent between the two patient groups.

The eleven CYP2D6 variants that were genotyped in iloperidone-treatedpatients are listed in Table 10, and their respective allele frequencyper race is provided in Table 11.

TABLE 11 CYP2D6 Allele Frequencies in Iloperidone-treated Patients Black& African Overall Asian Americans White Others Allele (N = 222) (N = 17)(N = 108) (N = 89) (N = 8) *2 39.2% (n = 174) 32.3% (n = 11) 44.4% (n =96) 34.3% (n = 61) 37.5% (n = 6)  *4^(†) 12.7% (n = 75) 12.0% (n = 6)11.0% (n = 33) 16.2% (n = 35) 3.8% (n = 1) *5 5.2% (n = 23) 2.9% (n = 1)6.0% (n = 13) 4.5% (n = 8) 6.3% (n = 1) *6 0.4% (n = 2) 0.0% (n = 0)0.5% (n = 1) 0.5% (n = 1) 0.0% (n = 0) *7 0.2% (n = 1) 2.9% (n = 1) 0.0%(n = 0) 0.0% (n = 0) 0.0% (n = 0) *8 0.0% (n = 0) 0.0% (n = 0) 0.0% (n =0) 0.0% (n = 0) 0.0% (n = 0) *9 1.8% (n = 8) 0.0% (n = 0) 0.5% (n = 1)3.9% (n = 7) 0.0% (n = 0) *10^(† ) 16.0% (n = 95) 20.0% (n = 10) 14.7%(n = 44) 17.1% (n = 37) 15.4% (n = 4) *17  9.5% (n = 42) 5.9% (n = 2)17.6% (n = 38) 1.1% (n = 2) 0.0% (n = 0) *41  5.9% (n = 26) 8.8% (n = 3)1.4% (n = 3) 11.2% (n = 20) 0.0% (n = 0) ^(†)Genotypes for an additional74 patients were obtained for markers *4 and *10, including 8 Asians, 42Black and African Americans, 19 Whites, and 5 from other racial groups.N and n denote the number of patients and the number of alleles,respectively, from which frequencies were determined.

Six non-functional CYP2D6 variants were genotyped: *3, *4, *5, *6, *7,and *8. The most common variant was *4, detected in 16.2%, 11.0%, and12.0% White, Black and African American, and Asian patients,respectively. It has been previously reported that the *4 variant wasthe most common non-functional CYP2D6 variant among Caucasians (˜20%)and African Americans (7.5%), while it was expected to be rare amongAsians (Bradford 2002). The *5 variant was observed at a frequency of3-6%, depending on the racial group, consistent with previous reports(Bradford 2002). As expected, *3, *6, *7 were rare, and *8 was notobserved in any patient.

Four variants were genotyped which are associated with reduced CYP2D6enzymatic activity: *10 has been observed frequently in Asia (38-70%),*17 has been reported in ˜22% of African Americans, *41 is believed tobe common among Caucasians (possibly ˜20%), and *9 has been observedonly in a small percentage of individuals (1-2%) (Bradford 2002).

The * 10 variant was observed in 15-20% of the iloperidone-treatedpatients across all racial groups. * 10 occurred more frequently thanexpected for Whites and African Americans, but less frequently amongAsians. Other studies have reported a high percentage of *10 in Asians(all above 38%) but a much lower percentage in Caucasians (4-8%) and inAfrican Americans (2-7%) (Bradford 2002). Variant *17 was the mostcommon variant in Black and African Americans (17.6%), and more rare inAsians (5.9%) and Whites (1.1%); this result is in agreement with theexpected frequencies for these populations (Bradford 2002).

As expected, the *41 variant was most common amongst Whites (11.2%) andrare in African Americans (1.4%). This variant, which has not beenextensively studied in other populations, was also seen in 8.8% ofAsians.

The functional *2 variant has been reported as the most commonlyoccurring variant coding for a CYP2D6 protein, with a slightly reducedactivity (˜80% of the wild type) (Bradford 2002). The *2 variant was themost commonly observed variant, with a frequency of 32 to 44% dependingof the racial group.

Because of the high frequency of the CYP2D6 variants, it is likely thata number of individuals carry more than one allele associated withreduced or abolished enzymatic activity.

Only 7 patients (3.1%) with 2 non-functional alleles were identified inthis study: 6 homozygotes *4/*4 and one compound heterozygote *5/*4.Seventy-two patients (32.1%) had only one functional copy of CYP2D6.Sixty-one patients (27.2%) had 2 functional copies of CYP2D6, with oneor 2 allelic variants with possible decreased enzymatic activity. Theother 84 patients (37.5%) carried only the *2 variant or were homozygotewt/wt at each variant locus. Linkage Disequilibrium (LD) analysisrevealed that several CYP2D6 loci were in complete linkagedisequilibrium. The *4 variant was in LD with * 10 (D′=1, LD 42.33) and*2 (D′=1, LD 5.75). The *2 variant was also in LD with *17 (D′=1, LD13.44), *10 (D′=1, LD 8.89), and *41 (D′=1, LD 5.43).

Analysis of individual CYP2D6 variants with iloperidone blood exposureshowed that the *4 and * 10 variants were significantly associated withthe (iloperidone+P88)/P95 ratio, with ratios of 2.28 for the *4 alleleas compared to 1.10 for the wt (p=2.8E-08) and 2.20 for the *10 alleleas compared to 1.03 for the wt (p=2.4E-09) (Table 12). Significantassociation was also seen for the *5 and *41 variants with ratios of2.16 for the *5 allele as compared to 1.19 for the wt (p=0.0016) and2.02 for the *41 allele as compared to 1.19 for the wt (p=0.0045) (Table12).

TABLE 12 Association of CYP2D6 Alleles With Exposure to Iloperidone atDay 14 Mean (Iloperidone + Variant Allele N P88)/P95 Ratio P valueCYP2D6 *2 *2 104 1.37 0.45 wt 60 1.21 CYP2D6 *4 *4 53 2.28 2.8E−08 wt165 1.10 CYP2D6 *5 *5 20 2.16 0.0016 wt 144 1.19 CYP2D6 *7 *7 1 1.970.61 wt 163 1.30 CYP2D6 *9 *9 8 1.33 0.96 wt 156 1.3 CYP2D6 *10 *10  672.20 2.4E−09 wt 151 1.03 CYP2D6 *17 *17  28 0.93 0.090 wt 136 1.39CYP2D6 *41 *41  23 2.02 0.0045 wt 141 1.19

As discussed previously, the presence of multiple CYP2D6 variants mayfurther affect the overall amount of CYP2D6 expressed and its totalenzymatic activity. Therefore, an additional analysis was conducted,taking into account the presence of all functional and non-functionalalleles as well as the expected decreased activity associated with someof the variants. A clear gradient of increased (iloperidone+P88)/P95ratio was observed with the presence of alleles associated withdecreased activity and non-functional alleles (Table 13). Patients with2 non-functional alleles were found to have an (iloperidone+P88)/P95ratio of 6.4, which was much higher than patients with only onenon-functional allele (1.8), with one or two alleles associated withdecreased activity (1.15), or with two functional alleles (0.80) (Table13).

TABLE 13 Effect of CYP2D6 Variants on Iloperidone Blood Exposure at Day14 Combination of 2 CYP2D6 Mean (Iloperidone + alleles N P88)/P95 RatioP value 2 functional alleles (*1 or 60 0.80 *2) 2 functional alleleswith 1 49 1.15 0.018 or 2 alleles associated with decreased activity 1non-functional allele, and 54 1.80 1.1E−07 1 functional alleleassociated or not with decreased activity 2 non-functional alleles 56.40 3.1E−17

Since a significantly decreased metabolism of iloperidone was observedin patients who carried at least one non-functional CYP2D6 allele,whether or not these patients had also an increased QTcF prolongationafter iloperidone treatment (Table 14) was also investigated. After 14days of treatment, patients with at least one non-functional CYP2D6allele had a significantly higher prolongation of the QTcF interval(16.3 msec) than those with 2 functional copies (9.7 msec, p=0.01). ByDay 28, the QTcF prolongation was reduced but was still statisticallydifferent between the 2 groups (11.4 and 4.4 msec respectively, p=0.02).

TABLE 14 Effect of CYP2D6 Variants on QTcF prolongation Day 14 Day 28QTcF change Mean (Iloperidone + QTcF change Mean (Iloperidone + CYP2D6alleles (msec) ^(†) P88)/P95 Ratio (msec) ^(†) P88)/P95 Ratio 2functional 9.7 (N = 110) 1.0 (N = 109) 4.4 (N = 90) 0.8 (N = 103)alleles 1 or 2 non- 16.3 (N = 59) 2.2 (N = 59) 11.4 (N = 56) 2.4 (N =59) functional alleles P = 0.01 P = 6.8E−08 P = 0.02 P = 1.4E−07 ^(†) LSSquares Mean QTcF change from baseline

D. Results and Discussion

Comparison based on the (iloperidone+P88)/P95 ratio regardless of thespecific CYP2D6 genotype revealed that patients with a ratio ≤1 have aQTc at Day 14 of 7.9 msec as compared to 16.0 msec for patients with aratio >1, p=0.0002 (Table 15). At Day 28, QTcF was reduced to 4.8 and10.1 msec for patients with a ratio ≤1 or >1 respectively.

TABLE 15 Effect of Iloperidone Blood Exposure on QTcF prolongation Mean(Iloperidone + Day 14 Day 28 P88)/P95 Ratio QTcF change (msec) ^(†) QTcFchange (msec) ^(†) ≤1 7.9 (N = 127) 4.8 (N = 119) >1 16.0 (N = 99) 10.1(N = 79) P = 0.0002 P = 0.062 ^(†) LS Squares Mean QTcF change frombaseline

The genotyping of multiple CYP2D6 variants in more than 200iloperidone-treated patients (Table 11) revealed that the CYP2D6*4 and*10 alleles, which are in linkage disequilibrium, were the most commonalleles associated with decreased or abolished enzymatic activity inWhites (16.2 and 17.1% respectively ) and Asians (12 and 20%respectively). In Black and African Americans, the * 17 allele was morecommon than the *4 and * 10 alleles (17.6% vs. 11 and 14.7%respectively). The frequency of the *5 allele was ˜5%, while the othernon-functional alleles were very rare. Most extensive analyses offrequency data of CYP2D6 variants came from European Caucasianspopulations, Chinese and Japanese populations, or selected Africanregions. To date, few studies have been reported on allele frequenciesin the US population, and some of the differences observed in this studywith data from European Caucasians, Africans, Chinese or Japanesepopulations are likely to reflect regional and national specificities ofUS populations.

It was observed that the *4, *10, *5 and *41 alleles were significantlyassociated with a reduced CYP2D6 iloperidone metabolism, morespecifically, an increase of the (iloperidone+P88)/P95 ratio (Table 12).When taking into account the genotype data of all alleles tested, thisratio was shown to be clearly dependent on the number of non-functionalalleles and of alleles associated with decreased activity (Table 13).

Furthermore it appears that the reduced iloperidone metabolism wasassociated with a higher QTcF prolongation after 14 and 28 days oftreatment. A significant difference was observed between patients withat least one-functional CYP2D6 allele and patients with 2 functionalalleles (Table 14). This difference was also observed between patientswith a (iloperidone+P88)/P95 ratio ≤1 or >1 regardless of their specificCYP2D6 genotypes (Table 15). These results offer a potential riskmanagement strategy and prospective testing tools for physicians whentreating patients with iloperidone if the potential for QTcFprolongation is considered to be a risk for the patient.

The starting point for determining the optimum dose of iloperidone is,as discussed above, a dose that has been shown to be acceptably safe andeffective in patients having a CYP2D6 genotype that results in a proteinhaving the same activity on iloperidone and P88 as the wild type CYP2D6protein. Such doses are known in the art and are disclosed, for example,in U.S. Pat. No. 5,364,866 discussed above.

Generally, the dose of iloperidone administered to a patient will bedecreased, as discussed above, if the enzymatic activity of the CYP2D6enzyme on iloperidone and P88 is less than about 75% of that of the wildtype CYP2D6. Enzymatic activity may be determined by any number ofmethods, including, for example, measuring the levels of iloperidoneand/or P88 in an individual's blood. In such a case, the iloperidonedose can be lowered such that measured levels of iloperidone and/or P88are substantially the same as levels measured in the blood ofindividuals having normal CYP2D6 enzymatic activity. For example, if theCYP2D6 enzymatic activity of a patient is estimated by one or moremethods (e.g., genotyping, determination of dextromorphan blood levels)to be 50% of the enzymatic activity normally observed in an individualhaving normal CYP2D6 enzymatic activity, the dose for the patient mayneed to be adjusted to one-half of the dose given to an individualhaving normal CYP2D6 enzymatic activity. Similarly, for ultrarapidmetabolizers, an analogous calculation will lead to the conclusion thata dose adjustment of twice that given an individual having normal CYP2D6enzymatic activity may be needed in order to achieve similar bloodlevels for the parent compound and active metabolites.

Alternatively, the dose of iloperidone administered to a patient may bedecreased based upon the patient's CYP2D6 genotype alone, or upon thepatient's P88:P95 or (iloperidone+P88):P95 ratios. For example, if apatient has a “poor metabolizer” genotype, or has a high P88:P95 or(iloperidone+P88):P95 ratio, the patient's dose of iloperidone may bereduced by, for example, 25%, 50%, or 75%. A patient's genotype can bereadily determined using standard techniques on samples of body fluidsor tissue. Such techniques are disclosed, e.g., in PCT ApplicationPublication Number WO03054226.

Furthermore, while the disclosure herein focuses on genotype, it isapparent to one of skill in the art that phenotype can also be used asan indicator of decreased activity of the CYP2D6 protein on iloperidoneand P88. For example, McElroy et al. describe a correlation betweenCYP2D6 phenotype and genotyping as determined bydextromethorphan/dextrorphan ratios. Therefore, although it is moreconvenient given the state of the art to look at genotype, if one wereto determine that a given patient expressed a mutant CYP2D6 with loweractivity on iloperidone and P88 than the wild type, or expressedabnormally low amounts of CYP2D6, then that patient would be given alower dose of iloperidone than a patient with wild type CYP2D6, asdiscussed above. Alternative methods for determining the relativeactivity of a patient's CYP2D6 gene include biochemical assays todirectly measure enzymatic activity, protein sequencing to examine theamino acid sequence of a patient's CYP2D6, monitoring transcription andtranslation levels, and sequencing the CYP2D6 gene mRNA transcript. Forexample, Chainuvati et al. describe assessment of the CYP2D6 phenotypeusing a multi-drug phenotyping cocktail (the Cooperstown 5+1 cocktail).

Iloperidone can be formulated into dosage units and administered topatients using techniques known in the art. See, e.g., PCT ApplicationPublication Number WO03054226, US Patent Application Publication Number20030091645, PCT Application Serial Number PCT EP03/07619, and PCTApplication Publication Number WO02064141, all of which are incorporatedherein by reference as though fully set forth.

In addition, the present invention provides a kit for determining apatient's CYP2D6 genotype and/or phenotype. Such a kit may include, forexample, a detection means, a collection device, containers, andinstructions, and may be used in determining a treatment strategy for apatient having one or more diseases or disorders for which iloperidonetreatment is indicated.

Detection means may detect a CYP2D6 polymorphism directly or may detectthe characteristic mRNA of the polymorphic gene or its polypeptideexpression product. In addition, as will be recognized by one of skillin the art, detection means may also detect polymorphisms in linkagedisequilibrium with a CYP2D6 polymorphism. Accordingly, any polymorphismin linkage disequilibrium with the CYP2D6 polymorphisms disclosed inthis application may be used to indirectly detect such a CYP2D6polymorphism, and is within the scope of the present invention.

Detection means suitable for use in the methods and devices of thepresent invention include those known in the art, such aspolynucleotides used in amplification, sequencing, and single nucleotidepolymorphism (SNP) detection techniques, Invader® assays (Third WaveTechnologies, Inc.), Taqman® assays (Applied Biosystems, Inc.), genechip assays (such as those available from Affymetrix, Inc. and RocheDiagnostics), pyrosequencing, fluorescence resonance energy transfer(FRET)-based cleavage assays, fluorescent polarization, denaturing highperformance liquid chromatography (DHPLC), mass spectrometry, andpolynucleotides having fluorescent or radiological tags used inamplification and sequencing.

A preferred embodiment of a kit of the present invention includes anInvader® assay, wherein a specific upstream “invader” oligonucleotideand a partially overlapping downstream probe together form a specificstructure when bound to a complementary DNA sequence. This structure isrecognized and cut at a specific site by the Cleavase enzyme, releasingthe 5′ flap of the probe oligonucleotide. This fragment then serves asthe “invader” oligonucleotide with respect to synthetic secondarytargets and secondary fluorescently-labeled signal probes contained in areaction mixture. This results in the specific cleavage of the secondarysignal probes by the Cleavase enzyme. Fluorescence signal is generatedwhen this secondary probe, labeled with dye molecules capable offluorescence resonance energy transfer, is cleaved. Cleavases havestringent requirements relative to the structure formed by theoverlapping DNA sequences or flaps and can, therefore, be used tospecifically detect single base pair mismatches immediately upstream ofthe cleavage site on the downstream DNA strand. See, e.g., Ryan et al.,Molecular Diagnosis, 4; 2:135-144 (1999); Lyamichev et al., NatureBiotechnology, 17:292-296 (1999); and U.S. Pat. Nos. 5,846,717 and6,001,567, both to Brow et al., all of which are hereby incorporatedherein by reference.

Another preferred embodiment of a kit of the present invention includesa detection means comprising at least one CYP2D6 genotypingoligonucleotide specific to alleles known to predict a patient'smetabolizer phenotype. More particularly, the means comprises anoligonucleotide specific for the CYP2D6G)846A or CYP2D6C100Tpolymorphism. The means may similarly comprise oligonucleotides specificfor each polymorphism as well as the wild type sequence.

Detection methods, means, and kits suitable for use in the presentinvention are described in International Publication Nos. WO 03/0544266and WO 03/038123, each of which is hereby incorporated herein byreference. It should also be understood that the methods of the presentinvention described herein generally may further comprise the use of akit according to the present invention.

Collection devices suitable for use in the present invention includedevices known in the art for collecting and/or storing a biologicalsample of an individual from which nucleic acids and/or polypeptides canbe isolated. Such biological samples include, for example, whole blood,semen, saliva, tears, urine, fecal material, sweat, buccal smears, skin,hair, and biopsy samples of organs and muscle. Accordingly, suitablecollection devices include, for example, specimen cups, swabs, glassslides, test tubes, lancets, and Vacutainer® tubes and kits.

The present invention encompasses treatment of a patient for any diseaseor condition that is ameliorated by administration of iloperidone. Asdiscussed above, such diseases or conditions include, for example,schizoaffective disorders including schizophrenia, depression includingbipolar depression, as well as other conditions such as cardiacarrythmias, Tourette's syndrome, psychotic disorders and delusionaldisorders.

A related aspect of the invention is a method for obtaining regulatoryapproval for a pharmaceutical composition comprising iloperidone or anactive metabolite thereof, or a pharmaceutically acceptable salt ofeither, which comprises including in proposed prescribing informationinstructions to determine whether or not a patient is a CYP2D6 poormetabolizer prior to determining what dose to administer to the patient.In another related aspect, the invention is a method for commercializing(i.e., selling and promoting) pharmaceutical compositions comprisingsuch compounds said method comprising obtaining regulatory approval ofthe composition by providing data to a regulatory agency demonstratingthat the composition is effective in treating humans when administeredin accordance with instructions to determine whether or not a patient isa CYP2D6 poor metabolizer prior to determining what dose to administerto the patient and then disseminating information concerning the use ofsuch composition in such manner to prescribers (e.g., physicians) orpatients or both.

Another aspect of the invention is a method for obtaining regulatoryapproval for the administration of iloperidone based, in part, onlabeling that instructs the administration of a lower dose if thepatient is already being administered a CYP2D6 inhibitor, e.g.,paroxetine, etc.

Embodiments

1. A method for treating a patient with an active pharmaceuticalingredient including at least one of: iloperidone, a pharmaceuticallyacceptable salt of iloperidone, an active metabolite of iloperidone, anda pharmaceutically acceptable salt of an active metabolite ofiloperidone, comprising the steps of: determining the patient's CYP2D6genotype; and administering to the patient an effective amount of theactive pharmaceutical ingredient, whereby the amount of the activepharmaceutical ingredient is determined based on the patient's CYP2D6genotype.

2. The method of embodiment 1, wherein the amount of the activepharmaceutical ingredient is decreased if the genotype indicatesdecreased enzymatic activity of the CYP2D6 enzyme relative to the wildtype.

3. The method of embodiment 2, wherein the amount of the activepharmaceutical ingredient is decreased if the patient's CYP2D6G1846Agenotype is AA.

4. The method of embodiment 2, wherein the amount of the activepharmaceutical ingredient is decreased if the patient's CYP2D6G1846Agenotype is GA.

5. The method of embodiment 2, wherein the amount of the activepharmaceutical ingredient is decreased if the patient's CYP2D6C100Tgenotype is TT.

6. The method of embodiment 2, wherein the amount of the activepharmaceutical ingredient is decreased if the patient's CYP2D6C100Tgenotype is CT.

7. The method of embodiment 2, wherein the amount of the activepharmaceutical ingredient is decreased if the patient's CYP2D6 genotypeis 100C>T; 1661G>C; 1846G>A.

8. The method of embodiment 2, wherein the amount of the activepharmaceutical ingredient is decreased if the patient's CYPD26 genotypeis 100C>T; 1661G>C; 4180G>C.

9. The method of embodiment 2, wherein the amount of the activepharmaceutical ingredient is decreased if the patient's CYP2D6 genotypeis CYP2D6 deleted.

10. The method of embodiment 2, wherein the amount of the activepharmaceutical ingredient is decreased if the patient's CYP2D6 genotypeis −1584C; −1235 A>G; −740 C>T;

678 G>A; CYP2D7 gene conversion in intron 1; 1661 G>C; 2850 C>T; 2988G>A; 4180 G>C.

11. The method of embodiment 1, wherein the patient is suffering from atleast one of schizophrenia, schizoaffective disorder, depression,bipolar mania/depression, cardiac arrhythmia, Tourette's Syndrome, apsychotic disorder, a delusional disorder, and schizophreniformdisorder.

12. The method of embodiment 11, wherein the patient is at risk for aprolonged QT interval.

13. A method for treating a patient who is a CYP2D6 poor metabolizerwith a pharmaceutically active ingredient including at least one of:iloperidone, a pharmaceutically acceptable salt of iloperidone, anactive metabolite of iloperidone, and a pharmaceutically acceptable saltof an active metabolite of iloperidone, wherein the patient isadministered a lower dosage than would be given to an individual who isnot a CYP2D6 poor metabolizer.

14. The method of embodiment 13, wherein the patient is determined to bea CYP2D6 poor metabolizer based on at least one of the patient'sgenotype, the patient's phenotype, and the fact that the patient isbeing treated with an agent that reduces CYP2D6 activity.

15. The method of embodiment 13, wherein the patient's genotype includesat least one CYP2D6 allele selected from a group consisting of 2549 Adeletion, 1846 G>A, 1707 T deletion, 2935 A>C, 1758 G>T, 2613-2615 AGAdeletion, 1023 C>T, 2850 C>T, 4180G>C, 1659 G>A, 1661 G>C, 2850 C>T,3183 G>A, −1584 C, −1235 A>G, −740C>T, −678 G>A, 100 C>T, 2988 G>A, andCYP2D6 deletion.

16. The method of embodiment 14, wherein the patient's genotype includesat least one deletion of the CYP2D6 gene.

17. The method of embodiment 15, wherein the patient's genotype includesa CYP2D7 gene conversion in intron 1.

18. The method of embodiment 13, wherein the patient is suffering fromat least one of schizophrenia, schizoaffective disorder, depression,bipolar mania/depression, cardiac arrhythmia, Tourette's Syndrome, apsychotic disorder, a delusional disorder, and schizophreniformdisorder.

19. A method of treating a patient with a pharmaceutically activeingredient including at least one of: iloperidone, a pharmaceuticallyacceptable salt of iloperidone, an active metabolite of iloperidone, anda pharmaceutically acceptable salt of an active metabolite ofiloperidone comprising the steps of: determining whether the patient isbeing administered a CYP2D6 inhibitor, and reducing the dosage of drugif the patient is being administered a CYP2D6 inhibitor.

20. The method of embodiment 19, wherein the CYP2D6 inhibitor includesat least one of paroxetine, dolasetron, venlafaxin, and fluoxetine.

21. The method of embodiment 19, wherein the patient is suffering fromat least one of schizophrenia, schizoaffective disorder, depression,bipolar mania/depression, cardiac arrhythmia, Tourette's Syndrome, apsychotic disorder, a delusional disorder, and schizophreniformdisorder.

22. A method for determining a patient's CYP2D6 phenotype comprising thesteps of: administering to the patient a quantity of at least one of:iloperidone, a pharmaceutically acceptable salt of iloperidone, anactive metabolite of iloperidone, and a pharmaceutically acceptable saltof an active metabolite of iloperidone; and determining a firstconcentration of at least one of iloperidone and an iloperidonemetabolite in the patient's blood.

23. The method of embodiment 22, wherein the iloperidone metabolite isselected from a group consisting of P88 and P95.

24. The method of embodiment 23, wherein a first concentration isdetermined for each of P88 and P95.

25. The method of embodiment 24, wherein the patient is designated apoor metabolizer if the ratio of first concentrations of P88 to P95 isgreater than or equal to about 2.0.

26. The method of embodiment 24, wherein the patient is designated apoor metabolizer if the ratio of first concentrations of(P88+iloperidone)/P95 is greater than or equal to about 1.0.

27. The method of embodiment 24, wherein the patient is designated apoor metabolizer if the ratio of first concentrations of iloperidone andP88 to P95 is greater than or equal to about 3.0.

28. The method of embodiment 22, further comprising the steps of:administering to the patient at least one CYP2D6 inhibitor; determininga second concentration of at least one of iloperidone and an iloperidonemetabolite in the patient's blood; and comparing the fuirst and secondconcentrations.

29. The method of embodiment 28, wherein the CYP2D6 inhibitor isselected from a group consisting of paroxetine, ketoconazole, andfluoxetine.

30. The method of embodiment 28, wherein a second concentration isdetermined for each of P88 and P95.

31. The method of embodiment 30, wherein the patient is designated apoor metabolizer if the ratio of second concentrations of P88 to P95 isgreater than or equal to about 2.0.

32. The method of embodiment 28, wherein a first and secondconcentration is determined for each of iloperidone, P88, and P95.

33. The method of embodiment 32, wherein the patient is designated apoor metabolizer if the ratio of second concentrations of iloperidoneand P88 to P95 is greater than or equal to about 3.0.

34. A method for determining whether a patient is at risk forprolongation of his or her QTc interval due to administration of atleast one of: iloperidone, a pharmaceutically acceptable salt ofiloperidone, an active metabolite of iloperidone, and a pharmaceuticallyacceptable salt of an active metabolite of iloperidone comprising thesteps of: measuring a first QTc interval; administering to the patient aquantity of at least one of: iloperidone, a pharmaceutically acceptablesalt of iloperidone, an active metabolite of iloperidone, and apharmaceutically acceptable salt of an active metabolite of iloperidone,measuring a second QTc interval; and comparing the first and second QTcinterval.

35. The method of embodiment 34, wherein the dose of iloperidoneadministered to the patient is about 24 milligrams per day.

36. The method of embodiment 34, further comprising the step ofadministering to the patient at least one CYP2D6 inhibitor after theadministering step.

37. The method of embodiment 36, wherein the CYP2D6 inhibitor isselected from a group consisting of paroxetine, ketoconazole, andfluoxetine.

While this invention has been described in conjunction with the specificembodiments outlined above, it is evident that many alternatives,modifications and variations will be apparent to those skilled in theart. Accordingly, the embodiments of the invention as set forth aboveare intended to be illustrative, not limiting. Various changes may bemade without departing from the spirit and scope of the invention asdefined in the following claims.

What is claimed is:
 1. A method for treating a patient with iloperidone,wherein the patient is suffering from schizophrenia, the methodcomprising the steps of: determining whether the patient demonstratesreduced CYP2D6-mediated metabolism relative to wild type by: obtainingor having obtained a biological sample from the patient; and performingor having performed a genotyping assay on the biological sample todetermine if the patient has a genotype associated with reducedCYP2D6-mediated metabolism; and if the patient has a CYP2D6 normalmetabolizer genotype, then internally administering iloperidone to thepatient in a first amount, and if the patient has a CYP2D6 poormetabolizer genotype, then internally administering iloperidone to thepatient in a second amount, wherein the first amount of iloperidonecauses an iloperidone blood exposure level that is therapeuticallyeffective in a patient having a CYP2D6 normal metabolizer genotype,wherein the second amount of iloperidone is one of 25%, 50%, or 75% ofthe first amount, and wherein a risk of QTc prolongation for a patienthaving a CYP2D6 poor metabolizer genotype is lower following theinternal administration of the second amount of iloperidone than itwould be if the iloperidone were administered in the first amount. 2.The method of claim 1, wherein the performing or having performed thegenotyping assay step comprises: extracting or having extracted genomicDNA or mRNA from the biological sample, and sequencing or havingsequenced CYP2D6 DNA derived from the extracted genomic DNA or from theextracted mRNA, wherein the sequencing or having sequenced step furthercomprises: amplifying or having amplified a CYP2D6 region in theextracted genomic DNA or mRNA to prepare a DNA sample enriched in DNAfrom the CYP2D6 gene region; and sequencing or having sequenced the DNAsample by hybridizing the DNA sample to nucleic acid probes to determineif the patient has a genotype associated with reduced CYP2D6-mediatedmetabolism.
 3. The method of claim 1, wherein the CYP2D6 poormetabolizer genotype includes two alleles, wherein each of the twoalleles is one of the following: *3, *4, *5, *6, *7, *8, *9, *10, *17,or *41, and wherein the second amount is 50% of the first amount.
 4. Themethod of claim 1, further comprising, if the patient has a CYP2D6intermediate metabolizer genotype, then internally administeringiloperidone to the patient in a third amount, wherein the third amountis greater than the second amount, and smaller than the first amount,and wherein a risk of QTc prolongation for a patient having a CYP2D6intermediate metabolizer genotype is lower following the internaladministration of the third amount of iloperidone than it would be ifthe iloperidone were administered in the first amount.
 5. The method ofclaim 4, wherein the CYP2D6 intermediate metabolizer genotype includestwo alleles, wherein one of the two alleles is one of the following: *3,*4, *5, *6, *7, *8, *9, *10, *17, or *41, wherein the other of the twoalleles is one of *1 or *2.
 6. The method of claim 1, wherein the firstamount of iloperidone is about 24 mg/day, and the second amount ofiloperidone is about 12 mg/day.
 7. The method of claim 4, wherein thefirst amount of iloperidone is about 24 mg/day, the second amount ofiloperidone is about 12 mg/day, and the third amount of iloperidone isat least 12 mg/day and smaller than 24 mg/day.
 8. The method of claim 1,wherein the first amount of iloperidone and the second amount ofiloperidone are provided in a controlled release depot formulation ofiloperidone.
 9. The method of claim 8, wherein the first amount ofiloperidone is up to 1000 mg, and wherein the second amount ofiloperidone is up to 500 mg.
 10. A method of treating a patient who issuffering from a schizoaffective disorder, depression, Tourette'ssyndrome, a psychotic disorder or a delusional disorder, the methodcomprising: determining if the patient has a genotype associated withreduced CYP2D6-mediated metabolism by obtaining or having obtained abiological sample from the patient, and performing or having performed agenotyping assay on the biological sample to determine whether thepatient has a genotype associated with reduced CYP2D6-mediatedmetabolism, and if the patient has a CYP2D6 normal metabolizer genotype,then internally administering iloperidone to the patient in a firstamount, and if the patient has a CYP2D6 poor metabolizer genotype, theninternally administering iloperidone to the patient in a second amount,wherein the first amount of iloperidone causes an iloperidone bloodexposure level that is therapeutically effective in a patient having aCYP2D6 normal metabolizer genotype, and wherein the second amount ofiloperidone is one of 25%, 50%, or 75% of first amount.
 11. The methodof claim 10, wherein the patient is at risk for a prolonged QT interval.12. The method of claim 10, wherein the CYP2D6 poor metabolizer genotypeincludes two alleles, wherein each of the two alleles is one of thefollowing: *3, *4, *5, *6, *7, *8, *9, *10, *17, or *41, and wherein thesecond amount is 50% of the first amount.
 13. The method of claim 10,further comprising, if the patient has a CYP2D6 intermediate metabolizergenotype, then internally administering iloperidone to the patient in athird amount, wherein the third amount is greater than the secondamount, and smaller than the first amount, and wherein a risk of QTcprolongation for a patient having a CYP2D6 intermediate metabolizergenotype is lower following the internal administration of the thirdamount of iloperidone than it would be if the iloperidone wereadministered in the first amount.
 14. The method of claim 13, whereinthe CYP2D6 intermediate metabolizer genotype includes two alleles,wherein one of the two alleles is one of the following: *3, *4, *5, *6,*7, *8, *9, *10, *17, or *41, wherein the other of the two alleles isone of *1 or *2.
 15. The method of claim 10, wherein the first amount ofiloperidone is about 24 mg/day, and the second amount of iloperidone is12 mg/day.
 16. The method of claim 10, wherein the first amount ofiloperidone is a controlled release depot formulation of iloperidoneincluding an amount of iloperidone of up to 1000 mg, and wherein thesecond amount of iloperidone is a controlled release depot formulationof iloperidone including an amount of iloperidone of up to about 500 mg.17. A method of treating a patient who is suffering from aschizoaffective disorder, depression, Tourette's syndrome, a psychoticdisorder or a delusional disorder, the method comprising: determining ifthe patient is at risk for iloperidone-induced QTc prolongation byobtaining or having obtained a biological sample from the patient, andperforming or having performed a genotyping assay on the biologicalsample to determine whether the patient has a CYP2D6 poor metabolizergenotype, wherein the presence of a CYP2D6 poor metabolizer genotypeindicates risk for iloperidone-induced QTc prolongation, and if thepatient is not at risk for iloperidone-induced QTc prolongation, theninternally administering iloperidone to the patient in a first amount,and if the patient is at risk for iloperidone-induced QTc prolongation,then internally administering iloperidone to the patient in a secondamount, wherein the first amount of iloperidone causes an iloperidoneblood exposure level that is therapeutically effective in a patient nothaving a CYP2D6 poor metabolizer genotype, and wherein the second amountof iloperidone is one of 25%, 50%, or 75% of first amount.
 18. Themethod of claim 17, wherein the patient is at risk foriloperidone-induced QTc prolongation, and is a CYP2D6 poor metabolizerhaving a CYP2D6 genotype including two alleles, wherein each of the twoalleles is one of the following: *3, *4, *5, *6, *7, *8, *9, *10, *17,or *41.
 19. The method of claim 17, wherein the first amount ofiloperidone is a controlled release depot formulation of iloperidoneincluding an amount of iloperidone of up to about 1000 mg, and whereinthe second amount of iloperidone is a controlled release depotformulation of iloperidone including an amount of iloperidone of up toabout 500 mg.
 20. The method of claim 17, wherein the method comprises:wherein the first amount of iloperidone is about 24 mg/day, and thesecond amount of iloperidone is about 12 mg/day.